US20060088519A1 - Malignant tumor-inhibiting preparation comprising Des A fibrin - Google Patents

Malignant tumor-inhibiting preparation comprising Des A fibrin Download PDF

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US20060088519A1
US20060088519A1 US11/314,443 US31444305A US2006088519A1 US 20060088519 A1 US20060088519 A1 US 20060088519A1 US 31444305 A US31444305 A US 31444305A US 2006088519 A1 US2006088519 A1 US 2006088519A1
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fibrin
malignant tumor
cells
des
fibrinogen
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Hirobumi Senga
Caixia Li
Yongling Wan
Lishui Chang
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Tobishi Pharmaceutical Co Ltd
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Tobishi Pharmaceutical Co Ltd
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Assigned to TOBISHI PHARMACEUTICAL CO., LTD. reassignment TOBISHI PHARMACEUTICAL CO., LTD. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: CHANG, LISHUI, LI, CAIXIA, SENGA, HIROBUMI, WAN, YONGLING
Publication of US20060088519A1 publication Critical patent/US20060088519A1/en
Priority to US11/818,783 priority Critical patent/US8481047B2/en
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/36—Blood coagulation or fibrinolysis factors
    • A61K38/363—Fibrinogen
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/04—Antineoplastic agents specific for metastasis

Definitions

  • the present invention relates to a malignant tumor-inhibiting preparation comprising Des A fibrin.
  • melanoma, lung cancer, liver cancer, pancreatic cancer and other malignant tumors with a high malignancy were difficult to detect in the early stages, so that by the time a malignant tumor is diagnosed, both the primary cancer and the metastatic cancer can be already existed simultaneously, and surgical treatment is impossible in many cases.
  • radiotherapies do not show good result for therapy of these malignant tumors.
  • most chemotherapeutic drugs in current clinical use such as adriamycin, work by directly attacking the malignant tumor cells, but since they simultaneously target normal cells, they have strong side-effects which are a problem for clinical use. Therefore, no revolutionary new drug has emerged in decades. In order to overcome such a situation, it looks forward to the new type drugs for treating malignant tumors.
  • Fibrin also called Des AB fibrin or fibrin II
  • FPA fibrinopeptide A
  • FB fibrinopeptide B
  • Batroxobin a thrombin-like serine protease produced from the venom of the snake Bothrops atrox moojeni , is a glycoprotein enzyme which releases only FPA from fibrinogen to produce Des A fibrin (also called fibrin I) (see for example Thromb. Haemost 36:9-13(1976) and Thromb. Diath. Haemorrh. 45(Suppl.):63-68(1971)).
  • the malignant tumor cell has characteristics of spreading and migration. “Spreading” of malignant tumor cells in this case, means that the round tumor cells form pseudopodia in response to some signal, and this is a biological behavior of tumor cells, which becomes the basis of tumor growth, invasion and metastasis.
  • “migration” of malignant tumor cells means that the tumor cells move from their original locations through repeated bindings and dissociations between ligands and cell adhesion molecules in the cell membrane in response to some signal, and this is also a biological behavior which becomes the basis of tumor invasion and metastasis.
  • Des A fibrin fibrinogen degradation product
  • a malignant tumor-inhibiting preparation comprising a novel active ingredient, which is a malignant tumor-inhibiting preparation.
  • Des A fibrin might have a different effect than fibrin on malignant tumor cells, and after exhaustive researches into the effect of Des A fibrin on malignant tumor cell spreading and migration, they discovered that Des A fibrin acts to inhibit the spreading and migration of malignant tumor cells.
  • the present invention is based on this finding.
  • the present invention relates to a malignant tumor-inhibiting preparation comprising Des A fibrin.
  • FIG. 1 is a photograph showing electrophoretogram of fibrinogen, Des A fibrin and fibrin.
  • FIG. 2 is a graph showing the electrophoretogram of FIG. 1 quantified by an image analyzing system.
  • FIG. 3 shows photomicrographs of melanoma cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and fibrin (F)-treated wells.
  • FIG. 4 is a graph of the spreading rates of melanoma cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and fibrin (F)-treated wells.
  • FIG. 5 shows photomicrographs of breast cancer cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and fibrin (F)-treated wells.
  • FIG. 6 is a graph of the spreading rates of breast cancer cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and fibrin (F)-treated wells.
  • FIG. 7 shows photomicrograph of fibrosarcoma cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and fibrin (F)-treated wells.
  • FIG. 8 is a graph of the spreading rates of fibrosarcoma cells which were cultured in the PBS (A), fibrinogen (B), batroxobin (C), thrombin (D), Des A fibrin (E) and (F) fibrin-treated wells.
  • FIG. 9 is a graph showing the effect of Des A fibrin amounts on spreading of melanoma cells.
  • the malignant tumor-inhibiting preparation of the present invention comprises Des A fibrin as an active ingredient.
  • Des A fibrin is a substance obtained by releasing FPA from fibrinogen.
  • Fibrinogen is a dimeric glycoprotein (A ⁇ B ⁇ ) 2 formed by the disulfide linkages of two subunits (A ⁇ B ⁇ ), each consisting of three chains of an A ⁇ chain, a B ⁇ chain and a ⁇ chain which are bound by disulfide linkages. Since the A ⁇ chain consists of 610 amino acids (68 kDa), the B ⁇ chain of 461 amino acids (54 kDa), and the ⁇ chain of 411 amino acids (48 kDa), the molecular weight of fibrinogen is 340 kDa (see Thromb. Res. 83:1-75(1996) and Blood Coagulation, Fibrinolysis and Kinin, Aoki, A. and Iwanaga, S.
  • FPA is a peptide corresponding to the 16 amino acids (NH 2 -Ala-Asp-Ser-Gly-Glu-Gly-Asp-Phe-Leu-Ala-Glu-Gly-Gly-Gly-Val-Arg) (Seq. ID No. 1) at the amino terminal end of the A ⁇ chain of fibrinogen.
  • Des A fibrin is the residue [( ⁇ B ⁇ ) 2 ] obtained when FPA is released from fibrinogen (see Thromb. Haemost. 36:9-13(1976) and Thromb. Diath. Haemorrh. 45 (Suppl.):63-68(1971)). Since the molecular weight of FPA is 1,536 Da, the molecular weight of Des A fibrin is calculated to be about 337 kDa.
  • thrombin-like enzymes can be used to release FPA from fibrinogen.
  • the thrombin-like enzyme is a kind of serine protease. Those derived from snake venom are commonly used. Specific examples include batroxobin (Tobishi Pharmaceutical Co., Ltd. and Beijing Tobishi Pharmaceutical Co., Ltd., Beijing, China, a subsidiary company of Tobishi Pharmaceutical Co., Ltd.), which is extracted and purified from the venom of Bothrops atrox moojeni , as well as ancrod and other thrombin-like enzymes (such as Crotalase) which are derived from snake venom and the like. These thrombin-like enzymes may be naturally occurring preparations or may be products of genetic recombination.
  • Des A fibrin can be manufactured for example by the methods described in (1) through (6) below using fibrinogen and a thrombin-like enzyme:
  • the fibrinogen and thrombin-like enzyme used to manufacture Des A fibrin in the present invention are themselves known substances, which can be easily obtained commercially or prepared.
  • Des A fibrin itself is also a known substance that can be prepared by the aforementioned methods.
  • malignant tumor-inhibiting preparation of the present invention is targeted at malignant tumors.
  • malignant tumors can be generally classified into epithelial malignant tumors and non-epithelial malignant tumors. About 90% of tumors are said to be epithelial tumors.
  • Non-epithelial malignant tumors can be further classified into malignant tumors derived from mesenchymal tissue, malignant tumors derived from neural tissue and malignant tumors derived from undifferentiated cells. Specific examples of each kind of malignant tumor are given below.
  • Adenocarcinomas (carcinomas derived from glandular epithelium, which occur throughout the body including the stomach, intestines, pancreas, trachea, lungs, mammary glands, ovaries, corpus uteri, prostate glands and the like, are supposed to constitute 70 to 80% of human cancers), squamous cell carcinomas (cancers derived from the stratified squamous epithelium and occurring in epithelial tissue of the epidermis, lips, tongue, throat, esophagus, anus, vulva, uterine cervix and the like, and pulmonary squamous epithelial cancers classified as non-small cell lung cancer), basal cell carcinomas (derived from basal cells of the skin and adnexa), transitional cell carcinomas (derived from transitional epithelium, such as bladder cancer), liver cell carcinomas (derived from hepatocytes), renal cell carcinomas (derived from renal epithelium), cholangiocarcinomas
  • Fibrosarcomas derived from connective tissue and fibrous tissue
  • liposarcomas derived from connective tissue and fatty tissue
  • chondrosarcomas derived from connective tissue and cartilaginous tissue
  • osteosarcomas derived from connective tissue and bone tissue
  • angiosarcomas derived from blood vessels
  • lymphangiosarcomas derived from lymphoducts
  • myelogenic leukemia derived from hemopoietic cells
  • monocytic leukemia derived from hemopoietic cells
  • malignant lymphoma derived from lymphoid tissue
  • lymphocytic leukemia derived from lymphoid tissue
  • plasmacytoma multiple myeloma, derived from lymphoid tissue
  • Hodgkin's cell derived from lymphoid tissue
  • leiomyosarcoma derived from smooth muscle
  • rhabdomyosarcoma derived from striated muscle
  • Neuroblastoma derived from neuroblasts
  • medulloblastoma derived from medulloblasts
  • malignant astrocytoma derived from astrocytes
  • retinoblastoma derived from retinoblasts
  • glioblastoma derived from glioblasts
  • malignant neurilenoma derived from Schwann cells
  • melanoma derived from neuroectoderm
  • Malignant teratoma derived from totipotent cells
  • nephroblastoma derived from nephroblasts
  • hepatoblastoma derived from hepatoblasts
  • mixed tumors derived from various types of cells.
  • the malignant tumor-inhibiting preparation of the present invention can be highly effective against epithelial malignant tumors, and against non-epithelial malignant tumors derived from neural tissue and mesenchymal tissue, particularly melanoma, breast cancer and fibrosarcoma.
  • the malignant tumor-inhibiting preparation of the present invention can inhibit tumor invasion and metastasis by inhibiting the spreading and migration of malignant tumor cells, and can thus inhibit malignant tumors.
  • “Spreading” of malignant tumor cells in this case means that the round tumor cells form pseudopodia in response to some signal, and this is a biological behavior of tumor cells, which becomes the basis of tumor cell proliferation, invasion and metastasis.
  • the “migration” of malignant tumor cells means that the movement of tumor cells from their original locations through repeated bindings and dissociations between ligands and cell adhesion molecules on the cell membrane in response to some signal, and this is also a biological behavior which becomes the basis of tumor invasion and metastasis.
  • the malignant tumor-inhibiting preparation of the present invention may comprise Des A fibrin either by itself or in combination with other active substances.
  • Examples of other active substances include antimetabolites such as fluorouracil, antitumor antibiotics such as adriamycin, alkylating agents such as dacarbazine, plant-derived anticancer drugs such as paclitaxel and the like.
  • any formulation in the Japanese Pharmacopoeia General Rules for Preparations can be applied to the formulation of the malignant tumor-inhibiting preparation of the present invention.
  • the formulation of the malignant tumor-inhibiting preparation of the present invention include injections for direct application inside the body (including suspensions and emulsions); ointments (including fatty ointments, emulsion ointments (creams), water-soluble ointments and the like), inhalants, liquids (including ophthalmic solutions, collunarium and the like), suppositories, patches, poultices, lotions and other external formulations; and internal formulations including tablets (including sugar-, film- and gelatin-coated), liquids, capsules, granules, powders (including grains), pills, syrups, troches and the like.
  • These formulations can be prepared by the methods described in the Japanese Pharmacopoeia General Rules for Preparations.
  • the malignant tumor-inhibiting preparation of the present invention may also include pharmacologically acceptable solid or liquid carriers or interventional therapy bases.
  • pharmacologically acceptable solid or liquid carriers include solvents, stabilizers, preservatives, solubilizing agents, emulsifiers, suspending agents, buffering agents, isotonizing agents, coloring agents, bases, thickeners, excipients, lubricants, binding agents, disintegrating agents, coating agents, corrigents and the like.
  • Specific examples include water, lactose, sucrose, fructose, glucose, mannitol, sorbitol and other sugars and sugar alcohols, crystalline cellulose, methylcellulose, ethylcellulose, hydroxypropylcellulose, low substituted hydroxypropylcellulose, hydroxypropylmethylcellulose, hydroxypropylmethylcellulose phthalate, hydroxypropylmethylcellulose acetate succinate, carmellose, carmellose calcium, carmellose sodium, croscarmellose sodium, carboxymethylethylcellulose, cellulose acetate phthalate and other celluloses and related derivatives, corn starch, wheat starch, rice starch, potato starch, dextrin, pregelatinized starch, partly pregelatinized starch, hydroxypropyl starch, sodium carboxymethyl starch, cyclodextrin, pullulan and other starches and related derivatives, agar, sodium alginate, acacia, gelatin, collagen, shellac, tragacanth,
  • intervention therapy bases include stents, artificial blood vessels and the like.
  • the amount of Des A fibrin contained in the malignant tumor-inhibiting preparation of the present invention varies depending on the formulation adopted.
  • Case examples include 0.01 to 900 mg per 1 g in the case of a formulation for internal use, 0.01 to 500 mg per 1 ml in the case of an injection or 0.01 to 500 mg per 1 g in the case of a formulation for external use.
  • the administered dose of the malignant tumor-inhibiting preparation of the present invention varies depending on the patient's weight, disease's property and condition, but is for example 0.1 to 5,000 mg or preferably 100 to 2,500 mg of Des A fibrin per day in the case of an adult.
  • Des A fibrin was prepared by using a thrombin-like enzyme to release FPA from fibrinogen. Moreover, production of Des A fibrin was identified through a comparison with fibrinogen and fibrin.
  • the thrombin-like enzyme batroxobin (Tobarpin®, Beijing Tobishi Pharmaceutical Co., Ltd., Beijing, China) was added to a phosphate-buffered (PBS) solution of human fibrinogen (F-4883, Sigma, MO, USA) to make a final reaction solution with fibrinogen concentration of 3.0 mg/ml and batroxobin concentration of 0.5 BU/ml, and incubated for 1 hour at 37° C. in order to prepare Des A fibrin.
  • PBS phosphate-buffered
  • a PBS solution of thrombin (Sigma, MO, USA) was added to a PBS solution of human fibrinogen (F-4883, Sigma, MO, USA) to make a final reaction solution with fibrinogen concentration of 3.0 mg/ml and thrombin concentration of 0.5 U/ml, and incubated for 1 hour at 37° C. in order to prepare fibrin.
  • a PBS solution of fibrinogen with a final concentration of 3.0 mg/ml was prepared using human fibrinogen (F-4883, Sigma, MO, USA) as an untreated control.
  • Des A fibrin in Process (1) mentioned above was identified by electrophoresis. Specifically, given that fibrinogen is reduced into three-type chains (A ⁇ , B ⁇ and ⁇ chains) by the following treatment, productions were evaluated by electrophoresis after reduction of Des A fibrin into three-type chains ( ⁇ , B ⁇ and ⁇ chains) and reduction of fibrin into three-type chains ( ⁇ , ⁇ and ⁇ chains).
  • the Des A fibrin obtained in Process (1) and the fibrin obtained in Process (2) were washed three times with sterilized isotonic sodium chloride solution, and boiled for 5 to 6 minutes in 0.5 ml of 2% SDS/2% beta-mercaptoethanol/5 M urea solution to break the disulfide bonds, and dissolved.
  • FIG. 1 The results are shown in FIG. 1 as an electrophoretogram.
  • Lane 1 and Lane 4 show fibrinogen
  • Lane 2 shows Des A fibrin
  • Lane 3 shows fibrin.
  • the lowest band represents the ⁇ chain
  • the second band from the bottom represents the B ⁇ chain
  • the third band from the bottom represents the A ⁇ chain.
  • the aforementioned electrophoretogram was quantified by using an image analyzing system (Furi Science & Technology Co., Ltd., Shanghai, China), and represented in FIG. 2 .
  • the left peak shows the band density of the A ⁇ chain
  • the middle peak shows the band density of the B ⁇ chain
  • the right peak shows the band density of the ⁇ chain.
  • Matrigel® (BD Biosciences, NJ, USA) was utilized to establish an artificial extracellular matrix environment in vitro, and three types of malignant tumor cells (melanoma, breast cancer and fibrosarcoma) were used to evaluate the effect of Des A fibrin on spreading of malignant tumor cells in this environment.
  • B16-BL6 mouse malignant melanoma cells (Academy of Chinese Medical Sciences, Beijing, China) as the melanoma cells were subcultured in RPMI-1640 medium (Gibco, MD, USA) containing 10% fetal bovine serum (FBS, HyClone, Utah, USA), and the cells were used in the present experiment.
  • MMT060562 mouse breast cancer cells (ATCC, VA, USA) as the breast cancer cells were subcultured in MEM medium (Gibco, MD, USA) containing 10% FBS and 1% nonessential amino acids solution (Non-Essential Amino Acids Solution, Gibco, MD, USA), and the cells were used in the present experiment.
  • HT-1080 human fibrosarcoma cells (Academy of Chinese Medical Sciences, Beijing, China) as the fibrosarcoma cells were subcultured in MEM medium (Gibco, MD, USA) containing 10% FBS, and the cells were used in the present experiment.
  • the BU bathroxobin unit
  • the BU bathroxobin unit
  • 2 BU being the activity to achieve coagulation in 19.0 ⁇ 0.2 seconds when 0.1 ml of batroxobin solution is added to 0.3 ml of standard human plasma containing citric acid at 37° C.
  • treatment E Des A fibrin
  • treatment F fibrin
  • FIG. 3 Photomicrographs (45 magnification) of the melanoma cells cultured for 2 hours in each different treated well, are shown in FIG. 3 .
  • Des A fibrin significantly inhibits the spreading of melanoma cells in the presence of Matrigel ( FIG. 4 , p ⁇ 0.01).
  • Des A fibrin effectively suppresses spreading of melanoma cells.
  • FIG. 5 Photomicrographs (45 magnification) of the breast cancer cells cultured for 2 hours in each different treated well, are shown in FIG. 5 .
  • Des A fibrin significantly inhibits the spreading of breast cancer cells in the presence of Matrigel ( FIG. 6 , p ⁇ 0.01).
  • Des A fibrin effectively suppresses spreading of breast cancer cells.
  • FIG. 7 Photomicrographs (45 magnification) of fibrosarcoma cells cultured for 2 hours in each different treated well, are shown in FIG. 7 .
  • Des A fibrin significantly inhibits the spreading of fibrosarcoma cells in the presence of Matrigel ( FIG. 7 , p ⁇ 0.01).
  • Des A fibrin effectively suppresses spreading of fibrosarcoma cells.
  • Des A fibrin produced by the action of batroxobin on fibrinogen is thought to increase as the amount of fibrinogen increases and as the reaction time between fibrinogen and batroxobin increases. Therefore, various amounts of Des A fibrin were prepared by reacting the same concentration of batroxobin (0.5 BU/ml) with differing concentrations of fibrinogen (0.1 mg/ml, 0.5 mg/ml, 1 mg/ml, 2 mg/ml, 3 mg/ml, 6 mg/ml, 9 mg/ml, 12 mg/ml and 15 mg/ml) for 30 seconds, 60 seconds and 90 seconds, respectively. The effects of which, on spreading of malignant tumor cells, were investigated. The experimental methods were similar to the Treatment E as mentioned in (2) The method of measuring cell spreading of Example 2 excluding the fibrinogen concentrations and reaction times. The results are shown in FIG. 9 .
  • Des A fibrin dose-dependently inhibits the spreading of melanoma cells.
  • B16-BL6 mouse malignant melanoma cells (Academy of Chinese Medical Sciences, Beijing, China) as the melanoma cells were subcultured in RPMI-1640 medium (Gibco, MD, USA) containing 10% fetal bovine serum (FBS, HyClone, Utah, USA), and the cells were used in the present experiment.
  • MMT060562 mouse breast cancer cells (ATCC, VA, USA) as the breast cancer cells were subcultured in MEM medium (Gibco, MD, USA) containing 10% FBS and 1% nonessential amino acids solution (Non-Essential Amino Acids Solution, Gibco, MD, USA), and the cells were used in the present experiment.
  • a cover glass was laid on each well of 6-well plates (Corning, NY, USA), and 2.5 ⁇ 10 4 B16-BL6 melanoma cells suspended in 2 ml of 10% FBS-containing RPMI-1640 medium or 2.5 ⁇ 10 4 MMT060562 breast cancer cells suspended in 2 ml of 10% FBS-containing MEM medium were seeded into the well and cultured for 24 hours in a CO 2 incubator.
  • Two milliliter of 0.05 mg/ml fibrinogen diluted with 10% FBS MEM or Des A fibrin (produced by addition of 0.05 mg/ml fibrinogen and 2 BU/ml batroxobin) was added to the breast cancer cultured wells, and it was incubated for 48 hours in a CO 2 incubator.
  • the Wright stained test is conducted, and the number of migrated cells to the scratch line were counted using a phase contrast microscope (Olympus, Tokyo, Japan) and represented as number of migrated cells per square millimeter of cover glass (number of cells/mm 2 ).
  • fibrinogen treatment was used as the control of Des A fibrin treatment.
  • Des A fibrin effectively inhibits the migration of melanoma cells.
  • the number of migrated cells (12 ⁇ 3 cells/mm 2 ) was significantly less than the number of migrated cells treated with fibrinogen (29 ⁇ 10 cells/mm 2 ) (p ⁇ 0.01).
  • Des A fibrin effectively inhibits the migration of breast cancer cells.
  • the malignant tumor-inhibiting preparation of the present invention can effectively inhibit the spreading and migration of malignant tumor cells. Therefore, the present invention can be advantageously used to inhibit malignant tumors.

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US6682520B2 (en) * 1999-08-23 2004-01-27 Bistech, Inc. Tissue volume reduction

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* Cited by examiner, † Cited by third party
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US20100247510A1 (en) * 2007-12-14 2010-09-30 Tobishi Pharmaceutical Co., Ltd. agent for reducing a side effect of an anticancer drug
RU2469740C2 (ru) * 2007-12-14 2012-12-20 Тобиси Фармасьютикал Ко., Лтд. Средство для снижения побочного эффекта противоракового лекарственного средства
US10624910B2 (en) * 2007-12-14 2020-04-21 Tobishi Pharmaceutical Co., Ltd. Thrombin-like enzyme for reducing a side effect of an anticancer drug
US20110165070A1 (en) * 2008-04-24 2011-07-07 The Australian National University Methods for radiolabeling synthetic polymers
US9381262B2 (en) * 2008-04-24 2016-07-05 The Australian National University Methods for radiolabeling synthetic polymers

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EP1762242B1 (fr) 2012-07-11
AU2005257508A1 (en) 2006-01-05
US8481047B2 (en) 2013-07-09
CA2566991A1 (fr) 2006-01-05
RU2358754C2 (ru) 2009-06-20
HK1100359A1 (en) 2007-09-21
CN1972706B (zh) 2012-03-21
JPWO2006001523A1 (ja) 2008-04-17
KR20070026846A (ko) 2007-03-08
CN1972706A (zh) 2007-05-30
EP1762242A1 (fr) 2007-03-14
HK1105091A1 (en) 2008-02-01
KR101032249B1 (ko) 2011-05-02
JP4716994B2 (ja) 2011-07-06
US20090018062A1 (en) 2009-01-15
RU2006145902A (ru) 2008-08-10
EP1762242A4 (fr) 2009-09-30
WO2006001523A1 (fr) 2006-01-05

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